The stereocontrolled synthesis of oligosaccharides based on sophisticated protection/deprotection, activation and coupling strategies has been well established. See, e.g., Danishefsky et al. J. Am. Chem. Soc., 111:6656 (1989); Okamoto et al., Tetrahedron, 46:5835 (1990); and Ito et al., Tetrahedron 46:89 (1990). A useful alternative to as the major planes have a problem in that their intrinsic desensitization due to dyes is great enzymes. Toone et al., Tetrahedron, 45:5365 (1989). One advantage of such enzymatic synthesis is the lack of extensive protection and deprotection steps. A disadvantage of such enzymatic synthesis is the apparent limitation of product formation that results from the specificity of glycosyltransferase and glycosidase enzymes.
Glycosyltransferases are highly specific enzymes that catalyze the transfer or activated donor monosaccharides to acceptor saccharides. That transfer results in the elongation or synthesis of an oligo- or polysaccharide.
A number of glycosyltransferase types have been described including sialyltransferases, N-acetylgalactosaminyltransferases, N-acetylglucos-aminyltransferases and the like. Beyer, et al., Adv. Enzymol., 52:23 (1981). The designation of those enzymes indicates the nature of the donor substrate. Thus, for example, a sialyltransferase transfers a sialic acid moiety to an acceptor molecule.
Within each of the general enzyme types set forth above, specific transferase enzymes are additionally designated by the type of glycosidic linkage formed. For example, a .beta.1,4-galactosyltransferase transfers a galactosyl moiety to an acceptor molecule, forming a .beta.1,4-glycosidic linkage with such acceptor.
Further, glycosyltransferases are characterized by the acceptor to which the donor glycosyl compound is transferred. A .beta.1,4-galactosyltransferase from bovine milk (GalT, EC 2.4.1.22) is known to accept N-acetylglucosamine (GlcNAc) and its glycosides (.beta.is better than .alpha.-glycoside) as acceptor substrates. See, e.g., Schanbacher, et al., J. Biol. Chem., 245:5057 (1970); Berliner, et al., Mol. Cell. Biochem., 62:37 (1984); Nunez, et al., Biochemistry, 19:495 (1980); Beyer, et al., Adv. Enzymol., 52:23 (1981); Barker, et al., J. Biol. Chem., 247:7135 (1972); and Babad, et al., J. Biol. Chem., 241:2672 (1966). Glucose and its .alpha.-and .beta.-glucosides are also acceptable; however, lactalbumin is required for .alpha.-glucosides. Beyer, et al., supra.
Taken together with the donor and linkage specificity set forth above, such acceptor specificity is used to define unique products of glycosyltransferase activity.
Oligosaccharides are considered to have a reducing end and a non-reducing end, whether or not the saccharide at the reducing end is in fact a reducing sugar. In accordance with accepted nomenclature, oligosaccharides are depicted herein with the non-reducing end on the left and the reducing end on the right.
All oligosaccharides described herein are, thus, described with the name or abbreviation for the non-reducing saccharide (i.e., Gal), followed by the configuration of the glycosidic bond (.alpha. or .beta.), the ring position of the non-reducing saccharide involved in the bond (1 or 2), the ring position of the reducing saccharide involved in the bond (2, 3, 4, 6 or 8), and then the name or abbreviation of the reducing saccharide (i.e., GlcNAc).
It is often extremely difficult to make synthetic saccharides that can be used to study naturally occurring synthetic routes by inhibiting the synthetic reactions. The lack of such synthetic inhibitors hampers attempts to investigate the effects of metabolic changes on carbohydrate production and turnover.
It is also often difficult to prepare novel, non-naturally occurring oligo- and polysaccharides that are useful as carriers or solubilizing agents for drugs and, which because of their non-natural structures, are resistant to degradation in vivo.
There is, therefore, a pressing need for oligosaccharide compounds and efficient methods of making the same that serve as substrates or inhibitors of transferase and glycosidase enzymes.